Functional Analysis of Semaphorin 5A In Vivo

Semaphorin 5A 体内功能分析

• 批准号: 7470078
• 负责人: ONANONG CHIVATAKARN
• 金额: $ 0.26万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-01 至 2008-08-16
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7470078
• 关键词: Address; Affinity; Animals; Antibodies; Arts; Axon; Binding; Biochemical; Biological; Biological Assay; Biological Neural Networks; Blocking Antibodies; CSPG3 gene; Cells; Chondroitin Sulfate Proteoglycan; Class; Complex; Cues; Defect; Development; Disease; Embryo; Embryonic Nervous System; Family; Family member; Fiber; Genes; Genetic; Goals; Growth; Heparan Sulfate Proteoglycan; Immune Sera; In Vitro; Injury; Inorganic Sulfates; Interruption; Knockout Mice; Language; Mediating; Membrane; Methods; Mission; Molecular; Mus; Muscle fasciculation; Nature; Nerve Regeneration; Nervous System Physiology; Nervous system structure; Neuraxis; Neurites; Neurologic; Neurons; Pattern; Personal Satisfaction; Play; Population; Proteins; Proteoglycan; Retinal Ganglion Cells; Role; Semaphorins; Signal Transduction; Synapses; Techniques; Testing; Unspecified or Sulfate Ion Sulfates; Visual system structure; Work; aggrecan; axon growth; axon guidance; cell growth; developmental disease; diencephalon; extracellular; in vivo; insight; interpeduncular nucleus; member; mutant; nervous system development; nestin protein; neural circuit; neuronal growth; phosphacan; protein distribution; receptor; repaired; research study; response; syndecan 3; versican

DESCRIPTION (provided by applicant): Proper nervous system functioning critically depends on the proper development of an intricate network of neuronal connectivity. It is well established that defects in neural network assembly or interruption of synaptic connections as a result of disease or injury leads to severe neurological deficits. The Semaphorins are one important class of molecules known to regulate axonal growth, guidance, and plasticity. We have identified Semaphorin 5A (Sema5A) as a bi-functional axon guidance molecule that regulates neuronal growth in a proteoglycan-dependent manner in vitro. Specifically, in the presence of chondroitin sulfate proteoglycans (CSPGs), Sema5A inhibits neurite outgrowth, whereas in the presence of heparan sulfate proteoglycans (HSPGs), SemaSA promotes neurite outgrowth. The studies outlined in this proposal directly build on our in vitro findings and are aimed at the functional characterization of Sema5A in specific neuronal populations during nervous system development in vivo. In specific Aim 1, I propose to conditionally ablate Sema5A in the embryonic nervous system in order to study its role during the development of a major limbic fiber tract called the fasciculus retroflexus. I propose to cross our Sema5A conditional mice with specific cre- driver lines to selectively ablate Sema5A in the embryonic nervous system. In specific Aim 2,1 will explore mechanistic questions to gain insights into how Sema5A regulates neuronal growth in a proteoglycan dependent manner. The methods I will use to achieve my goals include the use of state-of-the-art mouse genetics, immunohistochemical and biochemical techniques. To ask mechanistic questions related to Sema5A function, I will employ in vitro membrane stripe assays with primary neurons from wild-type and Sema5A mutant animals. As a whole, the studies proposed are anticipated to provide important insights into how Sema5A functions in the development of specific central nervous system fiber tracts, and importantly, reveal which proteoglycans switch Sema5A-mediated growth to Sema5A-mediated inhibition. If successful, our studies will provide the first mechanistic clues of how CSPGs influence neuronal growth. Lay language summary: Chondroitin sulfate proteoglycans (CSPGs) are large extracellular molecules that are increasingly being recognized as important regulators of nerve cell growth during development. In addition, CSPGs have been implicated in limiting nervous system regeneration following injury. The work ' proposed here is mission-oriented-it is aimed at understanding how CSPGs instruct nerve cells not to grow. We expect to uncover new biological principles that may be relevant for the treatment of developmental disorders and/or promote nervous system repair following injury or disease.

描述（由申请人提供）：神经系统的正常功能很大程度上取决于复杂的神经元连接网络的正确发展。众所周知，疾病或损伤导致的神经网络组装缺陷或突触连接中断会导致严重的神经缺陷。信号蛋白是一类重要的分子，已知可调节轴突生长、引导和可塑性。我们已经确定 Semaphorin 5A (Sema5A) 是一种双功能轴突引导分子，可在体外以蛋白聚糖依赖性方式调节神经元生长。具体来说，在硫酸软骨素蛋白多糖 (CSPG) 存在的情况下，Sema5A 抑制神经突生长，而在硫酸乙酰肝素蛋白多糖 (HSPG) 存在的情况下，SemaSA 促进神经突生长。本提案中概述的研究直接建立在我们的体外研究结果的基础上，旨在研究体内神经系统发育过程中特定神经元群中 Sema5A 的功能特征。在具体目标 1 中，我建议有条件地消融胚胎神经系统中的 Sema5A，以研究其在称为反屈束的主要边缘纤维束发育过程中的作用。我建议将我们的 Sema5A 条件小鼠与特定的 credriver 系杂交，以选择性地消除胚胎神经系统中的 Sema5A。具体目标 2,1 将探索机制问题，以深入了解 Sema5A 如何以蛋白聚糖依赖性方式调节神经元生长。我将用来实现我的目标的方法包括使用最先进的小鼠遗传学、免疫组织化学和生化技术。为了提出与 Sema5A 功能相关的机制问题，我将对野生型和 Sema5A 突变动物的原代神经元进行体外膜条纹测定。总的来说，所提出的研究预计将为 Sema5A 在特定中枢神经系统纤维束的发育中如何发挥作用提供重要见解，更重要的是，揭示哪些蛋白聚糖将 Sema5A 介导的生长转变为 Sema5A 介导的抑制。如果成功，我们的研究将为 CSPG 如何影响神经元生长提供第一个机制线索。通俗语言摘要：硫酸软骨素蛋白聚糖 (CSPG) 是一种大型细胞外分子，越来越多地被认为是发育过程中神经细胞生长的重要调节剂。此外，CSPG 还可能限制损伤后的神经系统再生。这里提出的这项工作是以任务为导向的——旨在了解 CSPG 如何指示神经细胞停止生长。我们期望发现可能与发育障碍的治疗和/或促进损伤或疾病后神经系统修复相关的新生物学原理。